Heretofore, as a process for preparing an optically active α-amino acid and an ester thereof by an enantio-selective hydrolysis reaction using a lipase, there has been disclosed a method in which, for example, only one of enantiomers of various kinds of amino acid esters is selectively hydrolyzed in water in the presence of a porcine pancreatic lipase, a lipase originated from Burkholderia cepacia (Pseudomonas cepacia), or a lipase originated from Rhizopus to obtain an optically active (S)-amino acid and an optically active (R)-amino acid ester (for example, see Non-Patent Literature 2).
However, according to this method, a large amount of an enzyme has been used, and there are problems that an E value which is an index of selectivity between enantiomers is generally low. When an optically active carboxylic acid which is a product is water-soluble, it is difficult to recover 100% of the product from the aqueous solution after completion of the reaction, and yet, in the presence of a large amount of water, lowering in optical purity occurs due to self-hydrolysis reaction of the substrate. Incidentally, the E value has widely been utilized as an index of selectivity of kinetic optical resolution (for example, see Non-Patent Literature 3.).
Also, as a conventional process for preparing an optically active α-amino acid and an ester thereof by an enantio-selective hydrolysis reaction using a protease, there is disclosed, for example, a method in which one of the enantiomers of tyrosine ethyl ester is selectively hydrolyzed in an acetonitrile-water mixed solvent in the presence of α-chymotrypsin, subtilisin Carlsberg and subtilisin BPN′ to obtain an optically active (S)-tyrosine and an optically active (R)-tyrosine ethyl ester (for example, see Non-Patent Literature 4). Here, various reactions were carried out by changing a water content in acetonitrile, and the most preferred results can be obtained in an amount of 5 to 10% (v/v) of water based on acetonitrile which is a solvent.
However, in the above-mentioned reaction systems, no hydrolysis is carried out with a system in which a water content is extremely little as 10 equivalent or less based on an amount of the substrate, nor referred to at all. In the system used in this case, an amount of water used as a substrate is still large, it is difficult to completely inhibit self-hydrolysis of the amino acid ester, and a substrate concentration based on the solvent is low so that it is not an industrially preferred method.    Non-Patent Literature 1: J. Med. Chem., 46, 4533 (2003)    Non-Patent Literature 2: Chirality, 8, 418 (1996)    Non-Patent Literature 3: J. Am. Chem. Soc., 104, 7294 (1982)    Non-Patent Literature 4: Biotechnology Letters, 13, (5), 317 (1991)    Non-Patent Literature 5: “Chemical Dictionary”, published by Tokyo Kagaku Dojin Co., Ltd., p. 948 (2000)    Patent Literature 1: WO 9706162    Patent Literature 2: WO 2004063198    Patent Literature 3: WO 2004084812    Patent Literature 4: WO 9803473    Patent Literature 5: WO 2005051304